Compositions and methods for dehydrated storage of on-board reagents in microfluidic devices

Abstract

Manufacturing methods and compositions are described for production of self-contained microfluidic cartridge devices with on-board reagents for molecular biological testing. Sensitive reagents are stored in dry form without lyophilization or freezing, and reconstituted at the point of use with either a biological sample or a sample eluate at the point of use. Manufacturing methods include sheet and roll fabrication processes where the reagents are printed in place and sealed within individual microfluidic cartridges before gel vitrification.

What is claimed is: 
     
       1. A microfluidic cartridge comprising on-board reagents for performing a molecular diagnostic assay, said microfluidic cartridge comprising a body with enclosed microfluidic works having at least one chamber or channel, said chamber or channel having an upstream fluid path joined to a sample inlet port and a downstream fluid path joined to a vent, wherein enclosed within said chamber or channel is at least one vitrified gel reagent spot, the vitrified gel reagent spot comprising:
 a) a DNA polymerase; 
 b) one or more excipients comprising a disaccharide or a trisaccharide; and 
 c) a fluorosurfactant. 
 
     
     
       2. The microfluidic cartridge of  claim 1 , wherein the DNA polymerase is a TAQ polymerase. 
     
     
       3. The microfluidic cartridge of  claim 1 , wherein the disaccharide is trehalose. 
     
     
       4. The microfluidic cartridge of  claim 1 , wherein the trisaccharide is melezitose or raffinose. 
     
     
       5. The microfluidic cartridge of  claim 1 , wherein the one or more excipients further comprises a high molecular weight polyethylene glycol. 
     
     
       6. The microfluidic cartridge of  claim 5 , wherein the high molecular weight polyethylene glycol has a molecular weight ranging from 0.1 to 5 MDa, inclusive. 
     
     
       7. The microfluidic cartridge of  claim 5 , wherein the high molecular weight polyethylene glycol is a linear polyethylene glycol. 
     
     
       8. The microfluidic cartridge of  claim 1 , wherein the one or more excipients further comprises an amino acid. 
     
     
       9. The microfluidic cartridge of  claim 1 , wherein the one or more excipients further comprises a carrier protein. 
     
     
       10. The microfluidic cartridge of  claim 9 , wherein the carrier protein is bovine serum albumin or fish gelatin. 
     
     
       11. The microfluidic cartridge of  claim 1 , wherein the fluorosurfactant is a non-ionic fluoroalkylsurfactant. 
     
     
       12. The microfluidic cartridge of  claim 1 , wherein the fluorosurfactant is Fluorosurfactant FC-4430. 
     
     
       13. The microfluidic cartridge of  claim 1 , further comprising a second gel reagent spot comprising a primer or primers. 
     
     
       14. The microfluidic cartridge of  claim 1 , further comprising one or more additional gel reagent spots, the one or more additional gel reagent spots independently comprising a probe or probes, a reverse transcriptase, an RNA polymerase, nucleotide triphosphates, a nucleic acid template, or a buffer. 
     
     
       15. The microfluidic cartridge of  claim 1 , wherein the microfluidic works comprises a plurality of chambers or channels fluidly connected as a parallel array of assay pathways between the sample inlet port and the vent, wherein within each assay pathway comprises at least one gel reagent spot therein. 
     
     
       16. The microfluidic cartridge of  claim 15 , wherein each assay pathway comprises a gel reagent spot comprising at least one unique primer pair. 
     
     
       17. The microfluidic cartridge of  claim 15 , wherein each assay pathway comprises a gel reagent spot comprising at least one probe. 
     
     
       18. The microfluidic cartridge of  claim 15 , wherein each assay pathway comprises a gel reagent spot comprising a DNA polymerase and a flourosurfactant. 
     
     
       19. The microfluidic cartridge of  claim 1 , wherein the vitrified gel reagent spot further comprises a PCR enhancer selected to form a glass at room temperature. 
     
     
       20. The microfluidic cartridge of  claim 19 , wherein the PCR enhancer is betaine, n-formyl morpholine, δ-valerolactam (2-piperidone), ε-caprolactam, 1,2-cyclopentanediol, PVP-10, PVP-40, or a mixture thereof. 
     
     
       21. The microfluidic cartridge of  claim 1 , wherein the vitrified gel reagent spot further comprises inulin, cellulose, derivatized cellulose, polyvinylpyrrolidone, lysine, arginine, or a Maillard reaction inhibitor. 
     
     
       22. A kit comprising the microfluidic cartridge of  claim 1  individually packaged in a gas tight pouch under a dry atmosphere with a desiccant, and comprising labeling and instructions for use.